I propose to study the physiology and the biochemistry of the mannosyl/glycosyl endocytosis receptor. This will include formulating a mechanism for the endocytic process that considers the following steps in some detail; cell surface binding and clustering, ligand release and delivery to the lysosome, receptor metabolism, and receptor recycling. Fluorescence spectroscopy will be developed as a method for determining the cell surface binding, and clustering of receptors so that the dynamics of these steps can be studied in a quantitative fashion. The role of ligand structure and other (non-receptor) membrane components in the cell surface steps will be evaluated. Specific antibodies to the receptor will be prepared and chemical methods for labeling the receptor in situ will be developed. Using these tools the intracellular pathway of ligand delivery to the lysosome will be studied. The contributions of receptor degradation, biosynthesis and recycling in the regulation of glycoprotein uptake will be determined. The specific endocytosis of lysosomal glycosidases is an effective potential mechanism for regulating their extracellular levels. I plan to extend this to include the lysosomal proteases. The role of alpha2-macroglobulin in the uptake of proteases will also be studied to determine if these two means of regulating extracellular protease levels are complimentary. The receptors role in the intracellular transport of the BANA hydrolases and lysosome biogenesis will be evaluated. I plan to study the specificity of the receptor for high mannose intermediates found in glycoprotein processing and correlate it with the carbohydrate composition of the BANA hydrolases and their intracellular location. Comparing this with the subcellular distribution of the receptor determined by similar techniques we will then use agents that alter the subcellular distribution of receptor to see if they affect the biosynthetic processing of the cathepsins.